Structure of the Chromophore Binding Pocket in the Pr State of Plant Phytochrome phyA
Structure of the Chromophore Binding Pocket in the Pr State of Plant Phytochrome phyA
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DOI：
10.1021/jp108265h
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发表时间：
2011-02-10
影响因子：
3.3
通讯作者：
Hildebrandt, Peter
中科院分区：
文献类型：
--
作者：
Mroginski, Maria Andrea;Kaminski, Steve;Hildebrandt, Peter
利用蓝藻光敏色素Cph 1（Cph 1 Delta 2）的感觉模块的晶体结构，建立了植物光敏色素phyA的同源结构模型。作为发色团，掺入天然植物色素移动素辅因子（Phi B）或藻蓝胆素（PCB）（Cphl中的天然辅因子）。通过分子动力学（MD）模拟进一步优化这些同源性模型，揭示了与晶体phyA结果的令人满意的总体一致性，所述晶体phyA结果来自小螺旋片段α（7）的重组，其导致远离辅因子的几个氨基酸的置换。残基的这种重新定位还包括天冬氨酸218，这样，代替其在Cph 1 Delta 2中的羰基功能，额外的水分子与环B和C NH基团形成氢键。为了验证phyA在发色团结合口袋中的结构模型，通过量子力学/分子力学（QM/MM）混合方法计算了辅因子的拉曼光谱，并与实验共振拉曼（RR）光谱进行了比较。计算和实验光谱之间的令人满意的整体协议被视为一个良好的质量的结构模型的指示。此外，次甲基桥拉伸模式和在所选位置的发色团的同位素标记的效果是非常好地再现，以允许确认甚至细节的次甲基桥的几何形状所预测的同源性模型。具体而言，它表明，实验RR谱是一致的环D相对于环C的扭转角，这是明显高于phyA-PCB（45度）和phyAP Φ B（42度）比Cph 1 Δ 2（30度）。从MD轨迹的不同点计算的拉曼光谱显示模式频率和强度的变化，反映了从快照到快照的结构波动。最低能量结构的快照光谱和所有快照光谱的总和提供了同样好的描述的实验数据。特别大的变化之间的快照注意到的吡咯环B和C，这反映了由水分子的波动在辅因子腔中所带来的氢键相互作用的改变的N-H面内弯曲模式。这种高估的水分子的流动性是目前的QM/MM方法的缺陷，由于缺乏适当的蛋白质力场，不能充分考虑在辅因子口袋中的静电的后果。
A homology structural model was generated for plant phytochrome phyA utilizing the crystal structure of the sensory module of cyanobacterial phytochrome Cphl (Cph1 Delta 2). As chromophores, either the native phytochromobilin cofactor (P Phi B) or phycocyanobilin (PCB), the natural cofactor in Cphl, was incorporated. These homology models were further optimized by molecular dynamics (MD) simulations revealing a satisfying overall agreement with the crystal phyA result from a restructuring of the small helical segment alpha(7) that leads to displacements of a few amino acids away from the cofactor. This repositioning of residues also include aspartate 218 such that, instead of its carbonyl function as in Cph1 Delta 2, an additional water molecule forms hydrogen bonds with the ring B and C NH groups. To validate the phyA structural model in the chromophore binding pocket, Raman spectra of the cofactor were calculated by means of the quantum mechanics/molecular mechanics (QM/MM) hybrid methodology and compared with the experimental resonance Raman (RR) spectra. The satisfactory overall agreement between calculated and experimental spectra is taken as an indication for the good quality of the structural model. Moreover, the methine bridge stretching modes and the effects of isotopic labeling at selected positions of the chromophore are very well reproduced to allow confirming even details of the methine bridge geometry as predicted by the homology model. Specifically, it is demonstrated that the experimental RR spectra are consistent with a torsional angle of ring D with respect to ring C that is distinctly higher for phyA-PCB (45 degrees) and phyA P Phi B (42 degrees) than for Cph1 Delta 2 (30 degrees). Raman spectra calculated from different points of the MD trajectory display variations of the mode frequencies and intensities reflecting the structural fluctuations from snapshot to snapshot. The snapshot spectrum of the lowest energy structure and the sum of all snapshot spectra afford an equally good description of the experimental data. Particularly large variations between the snapshots are noted for the N-H in-plane bending mode of the pyrrole rings B and C, which reflect alterations of the hydrogen bond interactions brought about by fluctuations of water molecules in the cofactor cavity. This overestimation of the water molecule mobility is a consequence of the deficiency of the current QM/MM methodology that, due to the lack of appropriate protein force fields, cannot adequately account for the electrostatics in the cofactor pocket.